【摘要】 蛋白磷酸酶在植物生长发育过程中发挥着重要的调控作用.酪氨酸蛋白磷酸酶属于蛋白磷酸酶中一个亚家族,在植物领域的研究中涉及较少.笔者所在实验室研究数据表明,酪氨酸蛋白磷酸酶19(PTP19)在植物盐响应中起负调控作用.使用MBP标签载体,构建MBP-PTP19融合蛋白表达载体,转化BL21大肠杆菌.以IPTG浓度、诱导时间、诱导温度3个因素设计L18正交实验探索MBP-PTP19蛋白的最佳诱导表达条件.找到该蛋白表达的最适诱导条件为IPTG终浓度0.1mmol/L、诱导时间4h、诱导温度37℃.该融合蛋白在上清液以及包涵体中均有分布.用直链淀粉树脂纯化得到了MBP-PTP19蛋白,并以其为抗原制备了PTP19的抗体,且抗体可识别PTP19蛋白,这为研究PTP19蛋白生化性质及其功能奠定了基础.更多还原

【Abstract】 Protein phosphotases play an important role in plant growth and development.Protein tyrosine phosphotases belongs to one subfamily of protein phosphotases,which were studied relatively less in the field of botany.Our preliminary study showed a protein tyrosine phosphatase 19(PTP19)in Arabidopsis plays a negative role in salt stress.In this study,construction of MBP-PTP19 fusion protein expression vector was performed with a vector containing MBP tag and transformed into Escherichia coli BL21.L18 orthogonal test was designed to explore appropriate expression condition of MBP-PTP19 protein through IPTG concentration,induction time,and induction temperature.It was found that the optimal inducing condition of PTP19 expression is induction with 0.1mmoL/L IPTG,at 37℃for 4h.The fusion protein was found in both supernatant and precipitate in bacteria.Amylose resin was used for PTP19 protein purification.The purified MBP-PTP19 fusion protein was used to prepare PTP19 antibodies,and the antibodies obtained could recognize PTP19 protein purified from bacteria.This will benefit the study of biochemical properties and functions of PTP19.更多还原